CD133 is well-documented to be expressed by tumor initiating cells and epithelial progenitor cells, which were proposed to have predominant roles for tumor recurrence and pre-metastatic niche formation, respectively. Thus, targeting CD133 might help eradicate the primary tumors and even prevent tumor metastasis. Herein, CD133-directed chimeric antigen receptor modified T cells (CART-133) were successfully generated and their marked antitumor activity was verified. Results from hematopoietic colony forming assays suggested that CART-133 cells may pose no irreversible myelosuppression. From October of 2015 to February of 2016, 10 patients with advanced and sorafenib-refractory hepatocellular carcinoma (HCC) were enrolled on phase I trial and were assigned into 3 dose-escalated cohorts (according to CART-133 positive cell amount: 0.10-1.0×106/kg, 2-5×106/kg, and 0.5-1.0×107/kg). 8 out of 10 patients received CART-133 monotherapy once or repeated infusion every 4-8 weeks. Patients who received CART-133 infusion were assessed for response. All patients had tolerable febrile syndromes during cell infusions. Of consecutively enrolled patients, rapid ascites growth occurred in 1 patient during infusion and was reversible by the use of diuretic, and 1 patient developed transiently drastic decline of hemoglobin and platelets and Grade 3 direct hyperbilirubinemia within 2 weeks after cell infusion. Reverse correlation between CD133+ cells in peripheral blood and CAR copy number in cohort 2 and 3 revealed an effective biological activity and safety of CART-133 and its rational expansion dose. 1 of 3 cases in cohort 1 aggressively progressed after cell therapy and became stable after transferred to cohort 2. Seven cases maintained stable disease as of the most recent follow-up, however, 2 patients died of upper gastrointestinal massive hemorrhage >9 weeks after infusion. On the basis of the presented data, additional 13 patients (5 sorafenib refractory HCCs, 5 advanced/metastatic pancreatic carcinomas, 2 metastatic colorectal carcinomas, and 1 advanced cholangiocarcinoma) were recruited into phase II trial using the expansion dose so far. All toxicities associated with the cell therapy even in those who received chemo-combined regimens with multiple cycle CART-133 infusions were controllable. The clinical response evaluation of all these patients in phase 2 is still ongoing. This study demonstrated the safety, feasibility, and preliminarily clinical efficacy of CART-133 treatment in epithelium-derived solid tumors, and guaranteed further patient recruitment. This clinical trial is registered at www.clinicaltrials.gov as NCT02541370.